The major goal is to understand the fundamental mechanisms for cellular commitment to mortality. We concentrate on the critical distinction between immortal early embryonic cells and mortal differentiating derivative cells. Our approach is to optimize the use of systematic genomic approaches for profiling gene expression patterns by large-scale cDNA sequencing and cDNA microarray technology with a current cohort of 15,257 genes collected from early mouse embryos. We are particularly focussing on four closely related experimental systems. (1) Differentiation of totipotent and immortal stem cells at E2.5 to mortal trophectoderm cells at E3.5. We have identified 16 novel genes that are dramatically up-regulated at the E3.5 blastocyst stage by comparing gene expression profiles between E2.5 morula stage and E3.5 blastocyst stage. To start to examine whether these genes might be involved in the immortal/mortal transition process, we are currently analyzing the nature of these genes in greater detail. (2) Differentiation and lineage commitment of pluripotent mouse embryonic stem (ES) cells. Expression profiling of mouse ES cells at six time points during the course of their differentiation has been done with the NIA mouse 15K cDNA microarrays. (3) Differentiation of the urogenital system: Germ line cells are often viewed as immortal, because they provide continuity from generation to generation and do not seem to age. To begin the analysis of gene expression regulation involved in germ line-somatic cell interactions, we conducted gene expression profiling by sequencing cell type-specific cDNA libraries that include E13.5 male and female primordial germ cells as well as two stages in the kidney-urogenital developmental axis, E12.5 female mesonephros and newborn ovary. We are currently analyzing genes differentially expressed in these cells, tissues, and organs more in detail. (4) Differentiation of hematopoietic stem cells: In an instance of particular interest, one of the genes (D7Wsu33e) that we previously isolated from E7.5 extraembryonic tissues turned out to be expressed essentially exclusively in hematopoietic cells. Accordingly the gene was renamed "Pan hematopoietic expression (Phemx)". The gene was strongly expressed in Lin- c-Kit+Sca-1+ hematopoietic stem cells, and expression was down-regulated after differentiation, but expression was still detected in all three hematopoietic lineages. We are currently analyzing the function of this gene.